Conventionally, there is an immunochromatography sensor as a typical example of a biosensor which is provided with a developing layer for developing a sample solution, includes a reagent part immobilized to a portion of the developing layer and a marked reagent part that is held by a portion of the developing layer in a dry state and is dissolvable by developing the sample solution, and measures the amount of the marker reagent bound to the reagent immobilization part, thereby to qualitatively or quantitatively analyze an analyte in the sample solution.
A general example of an immunochromatography sensor is provided with a sample applying part to which a sample solution is applied, and plural developing layers, and an antibody is immobilized to portions of the developing layers. Further, a marker antibody is held at the upper stream than the antibody immobilization part in a dry state so that it is dissolvable by the sample solution. When a required amount of sample solution is applied to the sample applying part, the sample solution penetrates through the developing layers, whereby measurement is started. A result of measurement is detected by the marker antibody that is bound to the antibody immobilization part. Particles of gold colloid are commonly used as a marker, and the binding to the antibody immobilization part is visually observable due to the particles of gold colloid. Thus, the result of measurement is obtained by visual observation. While sandwich reaction of antigen-antibody reaction is employed as a measurement principle, even when competition reaction is employed as a measurement principle, a result of measurement can be obtained by observing the state of binding of the marker reagent to the antibody immobilization part. In this specification, “immune chromatography” and “immunochromatography” denote the same chromatography, and it is an immunomeasurement method in which complexes of an immobilized reagent and a marker reagent are produced in a reaction layer comprising a wettable porous material, thereby to measure an analyte. That is, it is a measurement system utilizing antigen-antibody reaction. While the conventional immunomeasurement method needs a cleaning operation such as B/F separation, in the immunochromatography method, B/F separation is executed during the process in which the sample solution penetrates through a chromatography carrier as a reaction layer. Usually all reagents are in their dry states, and they are wetted by the sample solution during measurement. While gold colloid and latex are common as markers, magnetic particles, enzymes, and metal colloids other than gold colloid may be used. When the marker is an enzyme or the like, a user operation of adding an enzyme substrate or a reaction stopping agent is included as a measurement operation. Further, amongst the above-mentioned immunochromatography methods, one-step immunochromatography is a measurement method in which measurement is carried out by only a user operation of adding a sample solution. Since the fundamental measurement operation by the user is only application of a sample solution, it is called one-step immunochromatography. Further, although the above-described method requires qualitative judgement by visual observation, when a desired result of measurement is semi-quantitative or when judgement with accuracy higher than that is required, there is employed a method of reading a result of measurement by a transparent mode using an optical reading device, which is disclosed in Japanese Published Patent Application No. Hei.10-274624, or a method of capturing a result of measurement as an image with a camera or the like, and arithmetically processing the image, which is disclosed in Japanese Published Patent Application No. Hei.10-274653.
On the other hand, examples of a sensor device having the function of performing quantitative analyze by itself without requiring a measurement device for directly detecting a signal from the sensor by visual observation, have been disclosed in Japanese Patent No. 3005303, Japanese Published Patent Application No. Hei.7-159398, Japanese Published Patent Application No. Hei.8-278305. These inventions provide a sensor having the function of quantitative analysis by detecting the number of parts to which a marker reagent is bound among plural reagent immobilization parts, a sensor having the function of semi-quantitative analysis by varying the concentration in a reagent immobilization part, and a sensor which can simultaneously measure different target items in plural reagent immobilization areas.
In recent years, POCT (Point-of-Care Tests) is gradually becoming widespread in medical diagnosis scenes. In POCT, especially, a device that can measure an analyte speedily, easily, and precisely is desired. A fundamental principle employed for POCT has convenience that can deal with a wide range of analytes, and it is progressing in various fields not only clinical fields but also food hygiene fields, environmental measurement fields, and the like. On the other hand, although some POCT have quantitativeness for limited target items, most of POCT have only qualitative or semi-quantitative accuracy, and therefore, a technique that can measure an analyte more speedily, easily, and accurately and is applicable to wider fields has been demanded. However, while in the above-described method the analyte is measured by detecting the amount of the marker reagent bound to the reagent immobilization part in the sensor, the binding of the marker reagent to the reagent immobilization part has limitations. First of all, in the case of using sandwich reaction, a measurable antigen concentration area is eventually limited. Especially when it is antigen-antibody reaction, the antigen concentration in the area where the amount of binding linearly increases is about single or double digits. Even when more target antigen exists, it is saturated at a predetermined amount of binding, and the antigen exceeding the saturation level cannot be bound to the reagent immobilization part. When the target antigen further increases, a prozone phenomenon occurs. Therefore, when the concentration of the target antigen is high, previous dilution is needed. In order to perform dilution as well as execute highly precise quantitative analysis, dilution precision is also needed as a matter of course, and a device for dilution is required and, further, a dilution operation is required. Such dilution operation is extremely complicated for unskilled persons having little experience of chemical experiments, and therefore, the user must be selected. Furthermore, when such operational precision is not required, dilution can be carried out with relative ease by using a common pipette or the like. In this case, however, precision cannot be expected. Moreover, since the dilution operation is needed in addition to the measurement operation, extra time is required. Therefore, when speedy measurement in POCT is required, the measurement method using sandwich reaction can be used for only lower-accuracy qualitative analysis or semi-quantitative analysis. Further, a serious problem of the prozone phenomenon resides in that, even when the concentration of the actual analyte in the sample solution is high, a result apparently equivalent to a low concentration is undesirably obtained. For example, in the case of measurement in a clinical test, since a prescription for a patient is selected according to the test result, such prozone phenomenon might cause, in extreme cases, a problem relating to continuation of life. Accordingly, false-negative (FN) due to prozone phenomenon can be a fatal problem for the measurement.
Next, in the case of using competitive reaction, the amount of the marker reagent bound to the reagent immobilization part decreases with an increase in the concentration of the target antigen, and the marker reagent is not bound to the reagent immobilization part when the concentration of the target antigen is higher than a predetermined level. Also in this competitive reaction, when an antibody and an antigen are used as the immobilized reagent components, the target antigen concentration area is eventually limited due to the nature of binding, and a dilution operation is needed when the concentration of the target antigen is high, as in the above-mentioned sandwich reaction. In order to perform dilution as well as highly-precise quantitative analysis, dilution precision is also required as a matter of course, and a device for dilution is required and, furthermore, a dilution operation is required. Such dilution operation is extremely complicated for unskilled persons having little experience of chemical experiments, and therefore, the user must be selected. Furthermore, when such operational precision is not desired, dilution can be carried out with relative ease by using a common pipette or the like. In this case, however, precision cannot be expected. Moreover, since the dilution operation is needed in addition to the measurement operation, extra time is required. Therefore, when speedy measurement in POCT is required, the measurement method using competitive reaction can be used for only lower-accuracy qualitative analysis or semi-quantitative analysis. Further, only analytes having less change in target antigen concentration can be selected. Moreover, in order to measure an analyte having a wide concentration range without performing dilution, plural sensor devices must be used. When plural sensor devices are used, since the concentration of the analyte in the employed sample solution is not known by the operator, the operator must perform measurement twice, resulting in complicated workability and increased costs.